1. Field of the Invention
The present invention relates to a system and method for separating particles. The invention has particular advantages in connection with separating tumor cells from blood components, such as red blood cells and/or stem cells.
This application is related to U.S. Pat. No. 5,674,173, issued on Oct. 7, 1997; U.S. patent application Ser. No. 08/423,583, filed on Apr. 18, 1995 (abandoned); and U.S. patent application Ser. No. 08/634,167, filed on Apr. 18, 1996, U.S. Pat. No. 5,939,319. The entire disclosure of this U.S. patent and the entire disclosures of these U.S. patent applications are incorporated herein by reference.
2. Description of the Related Art
In many different fields, liquids carrying particle substances must be filtered or processed to obtain either a purified liquid or purified particle end product. In its broadest sense, a filter is any device capable of removing or separating particles from a substance. Thus, the term "filter" as used herein is not limited to a porous media material but includes many different types of processes where particles are either separated from one another or from liquid.
In the medical field, it is often necessary to filter blood. Whole blood consists of various liquid components and particle components. Sometimes, the particle components are referred to as "formed elements". The liquid portion of blood is largely made up of plasma, and the particle components primarily include red blood cells (erythrocytes), white blood cells (including leukocytes), and platelets (thrombocytes). While these constituents have similar densities, their average density relationship, in order of decreasing density, is as follows: red blood cells, white blood cells, platelets, and plasma. In addition, the particle constituents are related according to size, in order of decreasing size, as follows: white blood cells, red blood cells, and platelets.
In addition to red and white blood cells (and their subsets such as T cells and stem cells), plasma, and platelets, blood also includes in some cases, tumor cells. These cells have substantially similar densities, but different sedimentation velocities. Generally, stem cells have a sedimentation velocity greater than that of T cells and less than that of tumor cells.
Most current purification devices rely on density and size differences or surface chemistry characteristics to separate and/or filter blood components for transfusion or reinfusion purposes. Typically, blood components are separated or harvested from other blood components using a centrifuge. The centrifuge rotates a blood reservoir to separate components within the reservoir using centrifugal force. In use, blood enters the reservoir while it is rotating at a very rapid speed and centrifugal force stratifies the blood components, so that particular components may be separately removed. Although some centrifugal separation techniques are effective at separating some blood components from one another, many centrifugal separation processes are not capable of producing a highly purified end product.
In one type of separation procedure, a peripheral blood collection (withdrawn from an artery or vein) or a bone marrow blood collection is purified in a centrifugal separation process to isolate what is known as a peripheral cell collection or bone marrow cell collection, respectively. The peripheral cell collection or bone marrow cell collection primarily includes plasma, red blood cells, stem cells, and T cells, and may also include tumor cells if the donor's blood included such cells.
After undergoing a therapeutic treatment, such as chemotherapy or radiation therapy, to eliminate cancerous tumor cells, cancer patients often receive a transfusion of a peripheral cell collection or a bone marrow cell collection to replace stem cells destroyed as a side effect of the treatment. To reduce risks associated with infusing blood components from a foreign donor, some of these transfusions are autologous, where blood components collected from the patient are later reinfused back to the patient. However, the blood components initially collected from cancer patients may include cancerous tumor cells, which are then infused back into the cancer patient during reinfusion. This reinfusion of tumor cells may diminish the effectiveness of therapeutic treatments aimed at reducing cancerous tumor cells in a patient's body. Removal of tumor cells from a peripheral cell collection or bone marrow cell collection before transfusion, however, is extremely difficult.
Prior attempts to separate stem cells from tumor cells prior to reinfusion have had limited success. In one separation method, a selective antibody is introduced into a collected blood component sample after separating a substantial number of platelets and red blood cells from the sample. The selective antibody chemically attaches to stem cells to "mark" them. To separate the marked stem cells from the remaining blood components, the collected blood components are passed between stationary beads coated with a material which chemically bonds with the selective antibody. These chemical bonds retain the marked stem cells on the beads to filter them from the remaining blood components.
To remove the marked stem cells from the beads, an operator agitates the beads or flushes a chemical solution through the beads to break the chemical bonds between the material and selective antibody. This separation process, however, is extremely expensive, tedious, and time consuming. In addition, the beads do not remove a significant number of stem cells, and a substantial number of tumor cells often remain in the separated end product.
In another type of separation procedure, magnetic particles or fluid having an attached antibody are added to a blood component collection. The antibody chemically binds with stem cells in the sample to link the magnetic particles and stem cells together. To separate the stem cells, a magnetic separator is used to attract the linked magnetic substance and stem cells, and a substance is then added to break the chemical bonds between the stem cells and magnetic substance.
Although this separation procedure is capable of separating some stem cells and tumor cells, it is expensive and labor intensive. A significant number of tumor cells remain in the separated end product and a sizable number of stem cells are not recovered. In addition, the substances added to the blood sample in both the bead separation process and the magnetic separation process are potentially toxic if they are infused along with the separated blood components.
Centrifugal elutriation is another process used to separate tumor cells from stem cells. This process separates cells suspended in a liquid medium without the use of chemical antibodies. In one common form of elutriation, a cell batch is introduced into a flow of liquid elutriation buffer. This liquid which carries the cell batch in suspension, is then introduced into a funnel-shaped chamber located in a spinning centrifuge. As additional liquid buffer solution flows through the chamber, the liquid sweeps smaller sized, slower-sedimenting cells toward an elutriation boundary within the chamber, while larger, faster-sedimenting cells migrate to an area of the chamber having the greatest centrifugal force.
When the centrifugal force and force generated by the fluid flow are balanced, the fluid flow is increased to force slower-sedimenting cells from an exit port in the chamber, while faster-sedimenting cells are retained in the chamber. If fluid flow through the chamber is increased, progressively larger, faster-sedimenting cells may be removed from the chamber.
Thus, centrifugal elutriation separates particles having different sedimentation velocities. Stoke's law describes sedimentation velocity (SV) of a spherical particle, as follows: ##EQU1## where, r is the radius of the particle, .rho..sub.p is the density of the particle, .rho..sub.m is the density of the liquid medium, .eta. is the viscosity of the medium, and g is the gravitational or centrifugal acceleration. Because the radius of a particle is raised to the second power in the Stoke's equation and the density of the particle is not, the size of a cell, rather than its density, greatly influences its sedimentation rate. This explains why larger particles generally remain in a chamber during centrifugal elutriation, while smaller particles are released, if the particles have similar densities.
As described in U.S. Pat. No. 3,825,175 to Sartory, centrifugal elutriation has a number of limitations. In most of these processes, particles must be introduced within a flow of fluid medium in separate discontinuous batches to allow for sufficient particle separation. Thus, some elutriation processes only permit separation in particle batches and require an additional fluid medium to transport particles. In addition, flow forces must be precisely balanced against centrifugal force to allow for proper particle segregation.
Further, a Coriolis jetting effect takes place when liquid and particles flow into an elutriation chamber from a high centrifugal field toward a lower centrifugal field. The liquid and particles turbulently collide with an inner wall of the chamber facing the rotational direction of the centrifuge. This phenomenon mixes particles within the chamber and reduces the effectiveness of the separation process. Coriolis jetting also shunts flow of liquid and particles along the inner wall of the elutriation chamber from the inlet directly to the outlet. Thus, particles pass around the elutriative field to contaminate the end product.
If the combined density of particles and liquid in the elutriation chamber is significantly greater than the density of liquid entering the chamber, Coriolis jetting increases. This is because the relatively low density liquid entering the elutriation chamber encounters increased buoyant forces tending to accelerate the flow of liquid toward the outlet of the elutriation chamber. When the accelerated flow of liquid encounters tangential forces in the chamber, the flow of liquid may form a Coriolis jet capable of carrying larger, relatively faster sedimenting particles, such as tumor cells, around the elutriative field and through an outlet of the chamber.
Particle mixing by particle density inversion is an additional problem encountered in some prior elutriation processes. Fluid flowing within the elutriation chamber has a decreasing velocity as it flows in the centripetal direction from an entrance port toward an increased cross-sectional portion of the chamber. Because particles tend to concentrate within a flowing liquid in areas of lower flow velocity, rather than in areas of high flow velocity, the particles concentrate near the increased cross-sectional area of the chamber. Correspondingly, since flow velocity is greatest adjacent the entrance port, the particle concentration is reduced in this area. Density inversion of particles takes place when the centrifugal force urges the particles from the high particle concentration at the portion of increased cross-section toward the entrance port. This particle turnover reduces the effectiveness of particle separation by elutriation.
Particle adhesion is another problem associated with elutriation and other types of particle separation processes. Two types of particle adhesion reduce the effectiveness of particle separation. In the first type of particle adhesion, individual particles are bound to one another so that they act as groups of particles. When a substance including these particle groups is separated in a elutriative process or other separation process in which particle size is a factor, each group of particles acts as a larger particle and becomes separated along with larger particles, rather than being separated into smaller individual particles. In the second type of particle adhesion, particles adhere to equipment used during a separation procedure, such as plastic tubing or an elutriation chamber. This lowers the overall yield of particles, in particular, when separating a small number of particles.
For these and other reasons, there is a need to improve particle separation.